Catalase Enzyme Lab

Catalase Enzyme Data Tables

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Conclusion Questions

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Catalase Enzyme Lab Questions

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Catalase Lab Background

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Catalase Enzyme Protocol

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Enzyme Presentation

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BIG PICTURE

Enzyme = Catalase

Substrate = Hydrogen Peroxide (H2O2)

Catalase enzyme breaks down the substrate hydrogen peroxide into water and oxygen.

Variables Tested

Enzyme Concentration (i.e. How much Catalase is available)

Temperature

Substrate Concentration (i.e. How much Hydrogen Peroxide is around to break down?)

Ionic Concentration (i.e. How does the concentration of other things in the solution impact the reaction rate?)

 

 

Video of the Catalase Enzyme Lab Questions

BACKGROUND

Several enzymatic variables will be examined in this lab. You will be using the protein enzyme, catalase. Catalase is found in most cells, even in single-celled eukaryotes like yeast. In this lab, you will extract catalase from a yeast solution and test its catalytic effectiveness on hydrogen peroxide. Catalase speeds up the breakdown of peroxides which may form during respiration (metabolic energy production). This breakdown prevents the peroxide from causing unwanted oxidation of important biomolecules:

2 H2O2 → 2 H2O + O2 (gas)

We will measure enzyme activity by measuring the generation of oxygen gas — a product in the reaction.

Part A. The Time Course of Enzyme Activity

Procedure

  1. Set up the experimental apparatus as illustrated and described above.
  2. Obtain a reaction chamber.
  3. Obtain a bottle of 3% hydrogen peroxide (H2O2) solution and a 10mL graduated cylinder.
  4. Obtain a small amount of stock catalase (yeast) solution in a 50mL beaker. You will need 1.0mL of yeast solution for each trial. When you are ready, you will add it to the vial with a plastic pipette.
  5. Pour 10mL of hydrogen peroxide (H2O2) into the reaction chamber. Pipette in 1.0mL of stock catalase solution (yeast solution) and IMMEDIATELY stopper the reaction chamber tightly, submerge it in the water bath and place the plastic tubing into the bottom of the graduated cylinder, so all the bubbles formed in the reaction chamber are captured by the inverted graduated cylinder.
  6. Measure the gas levels in the graduated cylinder at 30-second intervals for 5 minutes. Record the levels in a data table of your own design.
  7. Plot the data on a graph. Don’t forget to label your axes and title your graph.

Part B. The Effect of Enzyme Concentration on Enzyme Activity

  1. Repeat the experiment from Part A, using 3 different levels of enzyme concentration: 75%, 50%, and 25% concentration of enzyme solution. You may easily do this by using the following procedures:
    a.    75% concentration: Follow the procedure from Part A, but use 0.75mL catalase solution in the reaction chamber, instead of 1.0mL.
    b.    50% concentration: Follow the procedure from Part A, but use 0.50mL catalase solution in the reaction chamber, instead of 1.0mL.
    c.    25% concentration: Follow the procedure from Part A, but use 0.25mL catalase solution in the reaction chamber, instead of 1.0mL.
  2. Record all data in a data table of your own design.
  3. Plot the data on the same graph as Part A. Don’t forget to clearly label the enzyme concentrations on your plotted lines.

Part C. The Effect of Temperature on Enzyme Activity

  1. Repeat the experiment from Part A (5 minute runs with 1mL catalase solution), using 3 different temperatures: 5°C, 37°C, and 100°C (boiled catalase). You may easily do this by using the following procedures:
    a.    5°C: Set up your reaction vessel and water bath and add ice to the water bath so that it is chilled to 5°C for 5 minutes before running the experiment. Keep adding ice to keep the temperature at 5°C or colder.
    b.    37°C: Set up your reaction vessel and water bath with heated water so that it is warmed to 37°C for 5 minutes before running the experiment. Keep adding hot water to keep the temperature at 37°C.
    c. 100°C (boiled catalase): We can’t use a water bath of boiling water since that may injure a student, so instead of keeping the reaction vessel in boiling water during the experiment, we will instead boil the catalase solution for 5 minutes. Then after the catalase solution has cooled you run the experiment in room temperature water (but with already boiled catalase).
  2. Record all data in a data table of your own design.
  3. Plot the data on a new graph. Also plot on this graph the room temperature data recorded in Part A. Don’t forget to clearly label your axes and plotted lines, and title your graph.

Part E. The Effect of Substrate Concentration on Enzyme Activity

  1. Repeat the experiment from Part A (5 minute runs with 1mL catalase solution), using 4 different substrate concentrations: 0%, 0.3%, 1.5%, and 3.0%. You may easily do this by using the following procedures
    a.    0%: Use 10mL distilled water only.
    b.    0.3%: Prepare this by adding 3mL of H2O2 to 7mL of distilled water.
    c.    1.5%: Prepare this by adding 5mL of H2O2 to 5mL of distilled water.
    d.    3.0%: This is the concentration of substrate from the original experiment in Part A; just use this initial data.
  2. Record all data in a data table of your own design.
  3. Plot the data on a new graph. Don’t forget to clearly label your axes and plotted lines, and title your graph.

Part F. The Effect of Ionic Concentration on Enzyme Activity

  1. Repeat the experiment from Part A (5 minute runs with 1mL catalase solution), using 3 different ionic concentrations: 10% NaCl, 2% NaCl, and 0% NaCl. You may easily do this by using the following procedures
    a.    10% NaCl: Make a 1.5% solution of H2O2 containing 10% NaCl by dissolving 5g of NaCl in 50mL of water then add 5mL of this solution to 5mL of H2O2.
    b.    2% NaCl: Make a 1.5% solution of H2O2 containing 2% NaCl by dissolving 1g of NaCl in 50mL of water then add 5mL of this solution to 5mL of H2O2.
    c.    0% NaCl: Prepare this by adding 5mL of distilled water to 5mL of H2O2.
  2. Record all data in a data table of your own design.
  3. Plot the data on a new graph. Don’t forget to clearly label your axes and plotted lines, and title your graph.

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